1. Field of the Invention
The present invention relates generally to the fields of cell biology, developmental biology and immunology. More particularly, it concerns methods and compositions relating to the identification, stimulation and repression of stems cells that express Toll-like receptors.
2. Description of Related Art
Survival depends on an innate immune system that can quickly recognize and respond to microbial/viral products. The Toll-like receptors (TLRs) are responsible for much of that recognition and consequently have vital roles (Takeda et al., 2003). Activation via TLRs couples innate immunity with the adaptive immunity provided by lymphocytes (Iwasaki and Medzhitov, 2004). For example, TLR ligands induce dendritic cells (DCs) to mature and support the differentiation of T helper 1 (Th1) cells. Cells responsible for both innate and adaptive immunity have finite lifespans and must be constantly replenished from hematopoietic stem cells (HSCs) and progenitors in bone marrow (Kondo et al., 2003). Although TLRs on mature immune cells have been well studied, little is known about when maturing cells in bone marrow acquire functional TLRs and whether those receptors influence hematopoietic development.
HSCs give rise to a series of progenitors that gradually lose differentiation options and produce cells of a given type. For example, multipotent progenitors (MPP) spawn common myeloid progenitors (CMP) that give rise to either megakaryocyte/erythrocyte progenitors (MEP) or granulocyte/macrophage progenitors (GMP) (Akashi et al., 2000). Early lymphoid progenitors (ELP) capable of producing T, B and NK cells give rise to pro-lymphocytes/common lymphoid progenitors (CLP) that can then become pre-B cells (Igarashi et al., 2002; Kouro et al., 2002; Kondo et al., 1997). All information available to date indicates that commitment to, and progression within, these lineages requires well studied growth and differentiation factors such as colony stimulating factors. These and other extracellular cues control expression of key transcription factors such as EBF, C/EBPα and PU.1 (Henderson and Calame, 1998; Rosmarin et al., 2005).
There is considerable controversy concerning the plasticity of stem/progenitors, and many studies have described experimental circumstances where cells of one hematopoietic lineage gave rise to cells of a different kind (Kondo et al., 2000; Iwasaki-Arai et al., 2003; Iwasaki et al., 2003; Xie et al., 2004). However, this is generally believed to represent latent differentiation potential that would not be utilized under physiological conditions.
The TLR family recognizes well conserved microbial/viral components. For example, TLR4 recognizes bacterial lipopolysaccharide (LPS) from Gram-negative bacteria (Hoshino et al., 1999) while TLR2 recognizes peptidoglycan and lipoteichoic acid from Gram-positive bacteria (Takeuchi et al,, 1999). Effective stimulation of cells via some TLRs requires cooperation with other molecules. The secreted MD-2 protein is associated with the extracellular portion of TLR4, and is essential for LPS recognition (Nagai et al., 2002). On B lymphocytes, the RP105/MD-1 complex cooperates with TLR2 and TLR4/MD-2 to cause antibody production to microbial membranes (Nagai et al., 2005). CD14 is known to cooperate with TLR2 and the TLR4/MD-2 complex in responses to lipoproteins and LPS respectively (Yoshimura et al., 1999; Means et al., 1999). In addition, TLRs require intracellular adaptor proteins for effective signaling. All TLRs except for TLR3 use the MyD88 adaptor protein for the production of inflammatory cytokines (Takeda and Akira, 2005). In addition, TLR3 and TLR4 use a MyD88-independent pathway, which is triggered by the TRIF/TICAM adaptor critical for induction of interferon-inducible genes (Yamamoto et al., 2003; Oshiumi et al., 2003).
A variety of defense mechanisms are triggered when microbial/viral products engage TLRs on innate immune cells. For example, TLR2/4 are linked to macrophage phagocytosis of bacteria (Blander and Medzhitov, 2004). TLR activation via MyD88 is required for phagosome maturation (Doyle et al., 2004). TLR signaling in DCs induces the expression of histocompatibility complex (MHC) and co-stimulatory molecules as well as the production of IL-12, a key cytokine for the induction of Th1 immune responses (Iwasaki and Medzhitov, 2004). In addition to these well-studied examples involving mature cells, there have been some hints that TLR might influence development within bone marrow. For example, chronic inflammation such as that elicited with endotoxin alters myeloid/lymphoid ratios in marrow (Ueda et al., 2004; Ueda et al,, 2005), and maturation of osteoclasts is altered by TLR ligands (Sato et al., 2004; Hayashi et al., 2003). Furthermore, Toll in Drosophila has a developmental role in determining dorso/ventral polarity and theoretically could contribute to other developmental processes (Ferrandon et al., 2004).